Novel chemokine for mobilizing stem cells

ABSTRACT

Novel chemokines for mobilizing stem cells are provided. Methods of mobilizing stem cells are also provided.

[0001] This application is a divisional of U.S. application Ser. No. 09/567,225, filed May 9, 2000, which is a continuation of U.S. application Ser. No. 09/225,501, filed Jan. 6, 1999 (now abandoned), which is a divisional of U.S. application Ser. No. 08/740,033, filed Oct. 23, 1996 (now abandoned), which claims the benefit of U.S. Provisional Application No. 60/006,051, filed Oct. 24, 1995. All of the above-listed applications are incorporated herein by reference.

BACKGROUND OF THE INVENTION

[0002] Hematopoietic cells have very important roles in a number of different processes in the body. For example, leukocytic hematopoietic cells are important in maintaining the body's defenses against disease; monocytes, macrophages and lymphocytes are involved in potentiating the body's responses to infection and tumors, while granulocytes are involved in overcoming infection, parasites and tumors. Platelets, another hematopoietic cell, form an important element in the hemostatic mechanism through initiating thrombus formation by their adhesion to each other and to damaged surfaces, and by the release of factors which assist in the formation of the fibrin clot. Erythrocytes are mainly involved in the transport of oxygen.

[0003] All of these blood cells are derived from a single progenitor cell called the hematopoietic stem cell. Stem cells are both pluripotent, in that they give rise to all different cell types, and capable of self renewal. Hematopoietic stem cells make up only a small percentage of bone marrow cells and are normally quiescent. However, when stimulated to divide, these stem cells produce a differentiated daughter cell with great proliferative potential. Sequential rounds of division and differentiation give rise to an enormous amplification of cell numbers which is necessary for the production of mature blood cells. This process of division and differentiation is subject to regulation at many levels to control cell production.

[0004] Numerous studies have led to the definition of functions of several hematopoietic regulatory messengers. These biomolecules have been characterized as stimulatory, e.g., Colony Stimulating Factors (CSFs) and interleukins (IL-1, IL-3, IL-5 and IL-9); inhibitory, e.g., transforming growth factor-β (TGF-β), interferon, prostaglandin E, tumor necrosis factor, macrophage inflammatory protein-1 (MIP-1), lactoferrin, acidic isoferritins, acSKDP, and pEEDCK (a synthetic HP5B monomer); or enhancing, e.g., TGF-β, IL-6, IL-4, IL-9, IL-11, MIP-1, MIP-2, leukemia inhibitory factor and Steel factor. Pelus et al., Experimental Hematology 22:239-247 (1994). Stimulatory biomolecules have been found to promote division of particular cell lineages. For example, G-CSF derives neutrophil production, while erythropoietin promotes formation of erythrocytes.

[0005] A number of these biomolecules and additional agents have been found to induce the mobilization of hematopoietic stem cells.

[0006] A single injection of IL-8 has been shown to induce mobilization of pluripotent stem cells that are able to provide permanent reconstitution of myeloid cells and of T and B lymphocytes. Laterveer et al., Blood 85:2269-2275 (1995). IL-8 belongs to a family of pro-inflammatory molecules called chemokines. This family has been divided into two subfamilies, the CXC and CC chemokines, based on whether the first two cysteine residues in a conserved motif are adjacent to each other or are separated by an intervening residue. In general, CXC, which include IL-8, melanoma growth-stimulating activity (MGSA) and platelet factor (PF4), are potent chemoattractants and activators of neutrophils but not monocytes. In contrast, CC chemokines, which include RANTES, monocyte chemotactic protein 1 (MCP-1) and MIP-1, are chemoattractants for monocytes but not neutrophils.

[0007] Stem cell inhibitors (SCIs) such as the CC chemokines, murine and human MIP-1α (LD78), have also been shown to enhance the release and mobilization of cells into the peripheral blood. WO 94/28916; Simm et al., Blood 84:2937 (1994).

[0008] Increased mobilization of stem cells in patients treated with sequentially administered interleukin-3 and GM-CSF compared with GM-CSF alone has been reported by Brugger et al., Blood 79:1193-1200 (1992). In addition, it has been shown that the absolute number of peripheral blood progenitor cells can be expanded in vitro by culture in a cocktail of cytokines, usually including SCF, IL-3, and either IL-6 or IL-1. Bodine, D., Experimental Hematology 23:293-295 (1995).

[0009] SK&F 107647, a hematoregulatory agent containing an ethylene bridge in place of the cysteine bridge of HP5B, has been demonstrated to be a potent simulator of in vitro myelopoiesis. Pelus et al., Experimental Hematology 22:239-247 (1994). Injection of SK&F 107647 in normal mice resulted in a two- to six-fold increase in serum colony-stimulating activity. Administration of this agent over four days resulted in significant increases in the number of granulocyte-macrophage, erythroid, and multipotential progenitor cells, as well as stimulating their cell cycle rates.

[0010] It has also been found that pretreatment with stem cell stimulating factor such as G-CSF can expand the pool of progenitor cells susceptible for mobilization by these agents, further increasing their mobilizing effect. For example, the combination of MIP-1 a with G-CSF was found to increase white cell count in the blood as compared to G-CSF alone. Simm et al., Blood 84:2937 (1994). Co-administration of SCI with G-CSF caused the enhanced mobilization of a number of cell types including neutrophils, monocytes, eosinphils, lymphocytes and basophil. WO 94/28916. Administration of G-CSF alone had no effect on the release of eosinphils or basophils after two days of administration. Similar effects were observed when other agents such as GM-CSF, f-Met-Leu-Phe or IL-8 were coadministered with SCIs.

[0011] New chemokines have now been identified which also mobilize stem cells in an animal. These chemokines can be administered alone, or in combination with a colony stimulating factor or hemoregulatory agent to enhance mobilization of stem cells.

SUMMARY OF THE INVENTION

[0012] An object of the present invention is to provide novel chemokines for the mobilization of stem cells in an animal.

[0013] Another object of the invention is to provide a method of mobilizing stem cells.

BRIEF DESCRIPTION OF THE DRAWINGS

[0014]FIG. 1 shows the sequence and alignment of the novel chemokines with known chemokines.

DETAILED DESCRIPTION OF THE INVENTION

[0015] In recent years, the availability of recombinant cytokines and the use of hematopoietic stem cell support have resulted in the widespread application of high-dose chemotherapy regimens designed to improve the success of cancer therapy. Despite significant advances, however, delayed recovery of hematopoiesis remains an important source of morbidity and mortality for patients treated with this approach. Since their discovery over 20 years ago, peripheral blood hematopoietic progenitor cells (PBPCs) have been increasingly used to supplement and even replace bone marrow as the source of hematopoietic support in a variety of situations.

[0016] Purified populations of cells are increasingly being used therapeutically and it would therefore be advantageous to be able to increase the number of circulating blood cells. It is useful to be able to harvest hematopoietic cells prior to chemotherapy or radiotherapy, thus, protecting them from harmful effects of this therapy; after therapy, the cells can be returned to the patient. It would therefore be highly beneficial to provide an agent which promoted the release and mobilization of a number of hematopoietic cells. Such an agent would be useful for enhancing the response to infection.

[0017] Peripheral blood cell transplantation is an important procedure in the treatment of cancer patients with high dose chemotherapy. In such treatment, patients are treated to induce clinical remission of their cancer, then during the remission, successive treatment with CSF, for example, by priming with cyclophosphamide then administration of G-CSF, causes eventual mobilization of cells from the bone marrow to the peripheral circulation for harvesting of leukophoresed blood; then the patient is given high dose chemotherapy or radiotherapy and the resultant bone marrow failure is compensated for by infusion of the stored blood or cells collected previously. This procedure may be modified by the omission of the initial induction of remission, and whole blood may be collected rather than leukophoresed blood. The mobilization effects of the present invention makes it a candidate both to replace CSFs in such cancer treatment regimes, and also to complement the mobilization effects of CSFs in combined treatments.

[0018] The two subfamilies of chemokines (CXC and CC) are ever-expanding, and presumably the individual members have similar, if slightly divergent, functions. The chemokines disclosed in the present invention are new members of the CC subfamily and are structurally similar to MPC-1, MPC-3, hRANTES, mMIP-1α, and mMIP-1β (FIG. 1). The effect of these chemokines in inducing leukophilia will find clinical and veterinary application in all utilities where the raising of hematopoietic cell levels is important. For example, a chemokine of the present invention can be used to enhance immune responses against chronic infections, particularly parasitic and bacterial infections. It may also have a role in promoting wound healing.

[0019] The chemoattractant activity of these chemokines can be boosted by pretreatment with a colony stimulating factor such as G-CSF or GM-CSF. Alternatively, the hematoregulatory peptides SK&F 107647 (currently in clinical trials), FLT-3 ligand (Immunex) or any other G-CSF mimetics (peptide and non-peptide) may be used. These stimulants may have an even more dramatic effect on these novel chemokines than on those already known due to their slight structural differences. For example, CKB-6 in combination with G-CSF was effective as a mobilizing factor. As known in the art, these peptides are useful in stimulating myelopoiesis in patients suffering from reduced myelopoietic activity, including bone marrow damage, agranulocytosis and aplastic anemia. Also included are patients who have depressed bone marrow function due to immunosuppressive treatment to suppress tissue reactions (i.e., bone marrow transplant surgery). They may also be used to promote more rapid regeneration of bone marrow after cytostatic chemotherapy and radiation therapy for neoplastic and viral diseases. There may also be a value where patients have serious infections due to a lack of immune response following bone marrow failure.

[0020] The hematopoietic stem cells released and harvested in the manner described above may be useful for subsequent in vitro and ex vivo manipulations to deliver gene products in gene therapy. Another embodiment is co-administration with cytotoxic drugs.

[0021] The following examples are provided for illustrative purposes only and are not intended to limit the invention.

EXAMPLES Example 1 Mobilization Assay for Novel Chemokines as Single Agents

[0022] A panel of novel chemokines will be tested as individual stem cell mobilization agents in BDF 1 mice. These chemokines include, but should not be limited to: Ckβ-1, Ckβ-4, Ckβ-6, Ckβ-7, Ckβ-8, Ckβ-9, Ckβ-10, Ckβ-11, Ckβ-12, Ckβ-13, and Ckα-1. Each agent will be assayed in concentrations of 50, 10, and 2 μg/mouse and administered via SC, IM, or a PO route. The kinetics of chemokine mobilization of stem cells will be monitored in 15 minute intervals over a period of 60 minutes by collecting blood samples from the mice by cardiac puncture. The mobilized stem cells will be collected by a densing gradient (Lympholyte M.). Cells are washed then frozen for future usage. The mobilization profile of the blood differentials will be assessed using a Technicon HI hematology analyzer. Mobilization of inflammatory cells such as PMNs, eosinophils, and basophils will be taken into account when evaluating the overall potential inflammatory profile. The chemokine IL-8, which mobilizes hematopoietic stem cells as a single factor, will be included in these studies as a positive control.

Example 2 Mobilization Assay for Novel Chemokines in Combination with Hematostimulants

[0023] J In these studies, hematostimulants will be assayed in combination with the aforementioned chemokines as mobilization factors. These agents include: G-CSF, GM-CSF, SK&F 107647, and FLT-3 ligand. However, any G-CSF mimetic (hematostimulants which are not colony stimulating factors like G-CSF or GMCSF, but have hematopoietic activity) may be used. In combination studies, G-CSF will be administered IP to mice four days prior to the novel chemokines. As in Example 1, the dose of chemokine and time of blood collection will be varied. Combination studies with hematostimulant pre-treatment will utilize MIP-1α as the positive control.

Example 3 CFU Assay

[0024] Blood samples collected during the mobilization phase will be assessed for colony forming units (CFU-GM) at days 7 and 14. Cells are adjusted to 2×10⁶ cells/ml in McCoys medium with 15×FBS serum. A single layer agar system utilizing the following is used: McCoys medium enriched with nutrients (NaHCO₃, pyruvate, amino acids and vitamins); 0.3% Bacto agar. To this is added cells from the blood samples (final concentration=2×10⁵ cells/ml). The agar plates are incubated at 37° C., 5% CO₂ for 7 days. Colonies of proliferating cells (CFU-GM) are counted utilizing a microscope. In addition, early hematopoietic high proliferative potential (HPP) progenitors, will be counted in the day 14 CFU cultures.

1 19 1 72 PRT CKBeta-1 1 Thr Lys Thr Glu Ser Ser Ser Arg Gly Pro Tyr His Pro Ser Glu Cys 1 5 10 15 Cys Phe Thr Tyr Thr Thr Tyr Lys Ile Pro Arg Gln Arg Ile Met Asp 20 25 30 Tyr Tyr Glu Thr Asn Ser Gln Cys Ser Lys Pro Gly Ile Val Phe Ile 35 40 45 Thr Lys Arg Gly His Ser Val Cys Thr Asn Pro Ser Asp Lys Trp Val 50 55 60 Gln Asp Tyr Ile Lys Asp Met Lys 65 70 2 70 PRT CKBeta-4 2 Ala Ser Asn Phe Asp Cys Cys Leu Gly Tyr Thr Asp Arg Ile Leu His 1 5 10 15 Pro Lys Phe Ile Val Gly Phe Thr Arg Gln Leu Ala Asn Glu Gly Cys 20 25 30 Asp Ile Asn Ala Ile Ile Phe His Thr Lys Lys Lys Leu Ser Val Cys 35 40 45 Ala Asn Pro Lys Gln Thr Trp Val Lys Tyr Ile Val Arg Leu Leu Ser 50 55 60 Lys Lys Val Lys Asn Met 65 70 3 70 PRT CKBeta-6 3 Val Val Ile Pro Ser Pro Cys Cys Met Phe Phe Val Ser Lys Arg Ile 1 5 10 15 Pro Glu Asn Arg Val Val Ser Tyr Gln Leu Ser Ser Arg Ser Thr Cys 20 25 30 Leu Lys Gly Gly Val Ile Phe Thr Thr Lys Lys Gly Gln Gln Phe Cys 35 40 45 Gly Asp Pro Lys Gln Glu Trp Val Gln Arg Tyr Met Lys Asn Leu Asp 50 55 60 Ala Lys Gln Lys Lys Ala 65 70 4 60 PRT CKBeta-7 4 Ala Gln Val Gly Thr Asn Lys Glu Leu Cys Cys Leu Val Tyr Thr Ser 1 5 10 15 Trp Gln Ile Pro Gln Lys Phe Ile Val Asp Tyr Ser Glu Thr Ser Pro 20 25 30 Gln Cys Pro Lys Pro Gly Val Ile Leu Leu Thr Lys Arg Gly Arg Gln 35 40 45 Ile Cys Ala Asp Pro Asn Lys Lys Trp Val Gln Lys 50 55 60 5 82 PRT CKBeta-8 5 Glu Asn Pro Val Leu Leu Asp Arg Phe His Ala Thr Ser Ala Asp Cys 1 5 10 15 Cys Ile Ser Tyr Thr Pro Arg Ser Ile Pro Cys Ser Leu Leu Glu Ser 20 25 30 Tyr Phe Glu Thr Asn Ser Glu Cys Ser Lys Pro Gly Val Ile Phe Leu 35 40 45 Thr Lys Lys Gly Arg Arg Phe Cys Ala Asn Pro Ser Asp Lys Gln Val 50 55 60 Gln Val Cys Met Arg Met Leu Lys Leu Asp Thr Arg Ile Lys Thr Arg 65 70 75 80 Lys Asn 6 79 PRT CKBeta-9 6 Ser Asp Ala Gly Gly Ala Gln Asp Cys Cys Leu Lys Tyr Ser Gln Arg 1 5 10 15 Lys Ile Pro Ala Lys Val Val Arg Ser Tyr Arg Lys Gln Glu Pro Ser 20 25 30 Leu Gly Cys Ser Ile Pro Ala Ile Leu Phe Leu Pro Arg Lys Arg Ser 35 40 45 Gln Ala Glu Leu Cys Ala Asp Pro Lys Glu Leu Trp Val Gln Gln Leu 50 55 60 Met Gln His Leu Asp Lys Thr Pro Ser Pro Gln Lys Pro Ala Gln 65 70 75 7 82 PRT CKBeta-10 7 Phe Asn Pro Gln Gly Leu Ala Gln Pro Asp Ala Leu Asn Val Pro Ser 1 5 10 15 Thr Cys Cys Phe Thr Phe Ser Ser Lys Lys Ile Ser Leu Gln Arg Leu 20 25 30 Lys Ser Tyr Val Ile Thr Thr Ser Arg Cys Pro Gln Lys Ala Val Ile 35 40 45 Phe Arg Thr Lys Leu Gly Lys Glu Ile Cys Ala Asp Pro Lys Glu Lys 50 55 60 Trp Val Gln Asn Tyr Met Lys His Leu Gly Arg Lys Ala His Thr Leu 65 70 75 80 Lys Thr 8 83 PRT CKBeta-11 8 Pro Ala Pro Thr Leu Ser Gly Thr Asn Asp Ala Glu Asp Cys Cys Leu 1 5 10 15 Ser Val Thr Gln Lys Pro Ile Pro Gly Tyr Ile Val Arg Asn Phe His 20 25 30 Tyr Leu Leu Ile Lys Asp Gly Cys Arg Val Pro Ala Val Val Phe Thr 35 40 45 Thr Leu Arg Gly Arg Gln Leu Cys Ala Pro Pro Asp Gln Pro Trp Val 50 55 60 Glu Arg Ile Ile Gln Arg Leu Gln Arg Thr Ser Ala Lys Met Lys Arg 65 70 75 80 Arg Ser Ser 9 82 PRT CKBeta-12 9 Arg Ser Gln Pro Lys Val Pro Glu Trp Val Asn Thr Pro Ser Thr Cys 1 5 10 15 Cys Leu Lys Tyr Tyr Glu Lys Val Leu Pro Arg Arg Leu Val Val Gly 20 25 30 Tyr Arg Lys Ala Leu Asn Cys His Leu Pro Ala Ile Ile Phe Val Thr 35 40 45 Lys Arg Asn Arg Glu Val Cys Thr Asn Pro Asn Asp Asp Trp Val Gln 50 55 60 Glu Tyr Ile Lys Asp Pro Asn Leu Pro Leu Leu Pro Thr Arg Asn Leu 65 70 75 80 Ser Thr 10 68 PRT CKBeta-13 10 Pro Tyr Gly Ala Asn Met Glu Asp Ser Val Cys Cys Arg Asp Tyr Val 1 5 10 15 Arg Tyr Arg Leu Pro Leu Arg Val Val Lys His Phe Tyr Trp Thr Ser 20 25 30 Asp Ser Cys Pro Arg Pro Gly Val Val Leu Leu Thr Phe Arg Asp Lys 35 40 45 Glu Ile Cys Ala Asp Pro Arg Val Pro Trp Val Lys Met Ile Leu Asn 50 55 60 Lys Leu Ser Gln 65 11 69 PRT hRANTES 11 Ala Ser Pro Tyr Ser Ser Asp Thr Thr Pro Cys Cys Phe Ala Tyr Ile 1 5 10 15 Ala Arg Pro Leu Pro Arg Ala His Ile Lys Glu Tyr Phe Tyr Thr Ser 20 25 30 Gly Lys Cys Ser Asn Pro Ala Val Val Phe Val Thr Arg Lys Asn Arg 35 40 45 Gln Val Cys Ala Asn Pro Glu Lys Lys Trp Val Arg Glu Tyr Ile Asn 50 55 60 Ser Leu Glu Met Ser 65 12 69 PRT mMIP-1alpha 12 Ala Pro Tyr Gly Ala Asp Thr Pro Thr Ala Cys Cys Phe Ser Tyr Ser 1 5 10 15 Arg Lys Ile Pro Arg Gln Phe Ile Val Asp Tyr Phe Glu Thr Ser Ser 20 25 30 Leu Cys Ser Gln Pro Gly Val Ile Phe Leu Thr Lys Arg Asn Arg Gln 35 40 45 Ile Cys Ala Asp Ser Lys Glu Thr Trp Val Gln Glu Tyr Ile Thr Asp 50 55 60 Leu Glu Leu Asn Ala 65 13 69 PRT mMIP-1beta 13 Ala Pro Met Gly Ser Asp Pro Pro Thr Ser Cys Cys Phe Ser Tyr Thr 1 5 10 15 Ser Arg Gln Leu His Arg Ser Phe Val Met Asp Tyr Tyr Glu Thr Ser 20 25 30 Ser Leu Cys Ser Lys Pro Ala Val Val Phe Leu Thr Lys Arg Gly Arg 35 40 45 Gln Ile Cys Ala Asn Pro Ser Glu Pro Trp Val Thr Glu Tyr Met Ser 50 55 60 Asp Leu Glu Leu Asn 65 14 78 PRT MCP-1 14 Leu Ala Gln Pro Asp Ala Ile Asn Ala Pro Val Thr Cys Cys Tyr Asn 1 5 10 15 Phe Thr Asn Arg Lys Ile Ser Val Gln Arg Leu Ala Ser Tyr Arg Arg 20 25 30 Ile Thr Ser Ser Lys Cys Pro Lys Glu Ala Val Ile Phe Lys Thr Ile 35 40 45 Val Ala Lys Glu Ile Cys Ala Asp Pro Lys Gln Lys Trp Val Gln Asp 50 55 60 Ser Met Asp His Leu Asp Lys Gln Thr Gln Thr Pro Lys Thr 65 70 75 15 82 PRT MCP-3 15 Ser Pro Gln Gly Leu Ala Gln Pro Val Gly Ile Asn Thr Ser Thr Thr 1 5 10 15 Cys Cys Tyr Arg Phe Ile Asn Lys Lys Ile Pro Lys Gln Arg Leu Glu 20 25 30 Ser Tyr Arg Arg Thr Thr Ser Ser His Cys Pro Arg Glu Ala Val Ile 35 40 45 Phe Lys Thr Lys Leu Asp Lys Glu Ile Cys Ala Asp Pro Thr Gln Lys 50 55 60 Trp Val Gln Asp Phe Met Lys His Leu Asp Lys Lys Thr Gln Thr Pro 65 70 75 80 Lys Leu 16 72 PRT NAP-1/IL-8 16 Ser Ala Lys Glu Leu Arg Cys Gln Cys Ile Lys Thr Tyr Ser Lys Pro 1 5 10 15 Phe His Pro Lys Phe Ile Lys Glu Leu Arg Val Ile Glu Ser Gly Pro 20 25 30 His Cys Ala Asn Thr Glu Ile Ile Val Lys Leu Ser Asp Gly Arg Glu 35 40 45 Leu Cys Leu Asp Pro Lys Glu Asn Trp Val Gln Arg Val Val Glu Lys 50 55 60 Phe Leu Lys Arg Ala Glu Asn Ser 65 70 17 71 PRT NAP-2 17 Ala Glu Leu Arg Cys Met Cys Ile Lys Thr Thr Ser Gly Ile His Pro 1 5 10 15 Lys Asn Ile Gln Ser Leu Glu Val Val Ile Gly Lys Gly Thr His Cys 20 25 30 Asn Gln Val Glu Val Ile Ala Thr Leu Lys Asp Gly Arg Lys Ile Cys 35 40 45 Leu Asp Pro Asp Ala Pro Arg Ile Lys Lys Ile Val Gln Lys Lys Leu 50 55 60 Ala Gly Asp Glu Ser Ala Asp 65 70 18 69 PRT hPF4 18 Glu Ala Glu Glu Asp Gly Asp Leu Gln Cys Leu Cys Val Lys Thr Thr 1 5 10 15 Ser Gln Val Arg Pro Arg His Ile Thr Ser Leu Glu Val Ile Lys Ala 20 25 30 Gly Pro His Cys Pro Thr Ala Gln Leu Ile Ala Thr Leu Lys Asn Gly 35 40 45 Arg Lys Ile Cys Leu Asp Leu Gln Ala Pro Leu Tyr Lys Lys Ile Leu 50 55 60 Lys Lys Leu Glu Ser 65 19 72 PRT CKApha-1 19 Val Leu Glu Val Tyr Tyr Thr Ser Leu Arg Cys Arg Cys Val Gln Glu 1 5 10 15 Ser Ser Val Phe Ile Pro Arg Arg Phe Ile Asp Arg Ile Gln Ile Leu 20 25 30 Pro Arg Gly Asn Gly Cys Pro Arg Lys Glu Ile Ile Val Trp Lys Lys 35 40 45 Asn Lys Ser Ile Val Cys Val Asp Pro Gln Ala Glu Trp Ile Gln Arg 50 55 60 Met Met Glu Val Leu Arg Lys Arg 65 70 

What is claimed is:
 1. A chemokine comprising the amino acid sequence of any one of SEQ ID NOS:1-10, wherein said chemokine is capable of mobilizing stem cells.
 2. A method of mobilizing stem cells in an animal comprising administering to an animal and effective amount of a chemokine comprising an amino acid sequence selected from the group consisting of any one of SEQ ID NOS: 1-10.
 3. The method of claim 2 further comprising administering a colony stimulating factor.
 4. The method of claim 2 further comprising administering a hematoregulatory agent. 